Malaria remains a major cause of human suffering with hundreds of million infections and many million deaths in the world annually. A major goal of research on human malaria is the development of vaccines targeting different stages of the parasite's life cycle in the human host and in the mosquito vector. The overall long term goal of our research is to develop a vaccine that would reduce or stop malaria transmission. This form of immunity, known as malaria transmission- blocking immunity, is largely antibody-mediated and operates in the mosquito midgut to prevent sexual reproduction of the parasites. A 27 kDa protein of Plasmodium falciparum (human malaria parasite) gametocytes has been identified as a target antigen for the development of transmission-blocking vaccine. Immunization in mice with various recombinant fragments of the 27 kDa protein elicited antibodies capable of suppressing malaria transmission. The linear B cell epitope recognized by transmission-blocking monoclonal antibodies was mapped using recombinant overlapping fragments of the 27 kDa protein expressed in E. coli and synthetic peptides. To facilitate rational vaccine development, we have also mapped, in addition to the B cell epitope, several helper T cell epitopes recognized by the 27 kDa protein- specific murine and human T cell clones. It now appears feasible to combine the mapped B and T cell epitopes to develop a subunit vaccine construct. A major goal of the studies proposed in this application is to investigate the biological mechanism and molecular basis of malaria transmission-blocking mediated by monoclonal antibodies or antibodies induced by the recombinant 27 kDa protein. Studies are also proposed to clone the 27 kDa protein homologue in an evolutionarily-related parasite P. gallinaceum which will offer an opportunity for vaccine trials using an animal malaria model system. Finally, studies are also proposed to construct a hybrid of two target antigens of transmission- blocking immunity: the 27 kDa gametocyte protein and the 25 kDa zygote/ookinete protein by recombinant cloning in Pichia pastoris and in DNA vaccine vectors. Evaluation of these two antigens either combined individually or as a hybrid molecule would identify an effective vaccine construct. Overall, the proposed studies would lead to an improved biological understanding of the sexual stages of human malaria parasite and suggest ways to produce an effective malaria transmission-blocking vaccine.